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Fri, 16 Jan 2015 14:51:48 +0100Fri, 16 Jan 2015 14:51:48 +0100Selective Lentiviral Gene Delivery to CD133-Expressing Human Glioblastoma Stem Cellshttp://publikationen.ub.uni-frankfurt.de/frontdoor/index/index/docId/36436
Glioblastoma multiforme (GBM) is a deadly primary brain malignancy. Glioblastoma stem cells (GSC), which have the ability to self-renew and differentiate into tumor lineages, are believed to cause tumor recurrence due to their resistance to current therapies. A subset of GSCs is marked by cell surface expression of CD133, a glycosylated pentaspan transmembrane protein. The study of CD133-expressing GSCs has been limited by the relative paucity of genetic tools that specifically target them. Here, we present CD133-LV, a lentiviral vector presenting a single chain antibody against CD133 on its envelope, as a vehicle for the selective transduction of CD133-expressing GSCs. We show that CD133-LV selectively transduces CD133+ human GSCs in dose-dependent manner and that transduced cells maintain their stem-like properties. The transduction efficiency of CD133-LV is reduced by an antibody that recognizes the same epitope on CD133 as the viral envelope and by shRNA-mediated knockdown of CD133. Conversely, the rate of transduction by CD133-LV is augmented by overexpression of CD133 in primary human GBM cultures. CD133-LV selectively transduces CD133-expressing cells in intracranial human GBM xenografts in NOD.SCID mice, but spares normal mouse brain tissue, neurons derived from human embryonic stem cells and primary human astrocytes. Our findings indicate that CD133-LV represents a novel tool for the selective genetic manipulation of CD133-expressing GSCs, and can be used to answer important questions about how these cells contribute to tumor biology and therapy resistance.N. Sumru Bayin; Aram S. Modrek; August Dietrich; Jonathan Lebowitz; Tobias Abel; Hae-Ri Song; Markus Schober; David Zagzag; Christian J. Buchholz; Moses V. Chao; Dimitris G. Placantonakisarticlehttp://publikationen.ub.uni-frankfurt.de/frontdoor/index/index/docId/36436Fri, 16 Jan 2015 14:51:48 +0100Measles virus glycoprotein-based lentiviral targeting vectors that avoid neutralizing antibodieshttp://publikationen.ub.uni-frankfurt.de/frontdoor/index/index/docId/26759
Lentiviral vectors (LVs) are potent gene transfer vehicles frequently applied in research and recently also in clinical trials. Retargeting LV entry to cell types of interest is a key issue to improve gene transfer safety and efficacy. Recently, we have developed a targeting method for LVs by incorporating engineered measles virus (MV) glycoproteins, the hemagglutinin (H), responsible for receptor recognition, and the fusion protein into their envelope. The H protein displays a single-chain antibody (scFv) specific for the target receptor and is ablated for recognition of the MV receptors CD46 and SLAM by point mutations in its ectodomain. A potential hindrance to systemic administration in humans is pre-existing MV-specific immunity due to vaccination or natural infection. We compared transduction of targeting vectors and non-targeting vectors pseudotyped with MV glycoproteins unmodified in their ectodomains (MV-LV) in presence of α-MV antibody-positive human plasma. At plasma dilution 1:160 MV-LV was almost completely neutralized, whereas targeting vectors showed relative transduction efficiencies from 60% to 90%. Furthermore, at plasma dilution 1:80 an at least 4-times higher multiplicity of infection (MOI) of MV-LV had to be applied to obtain similar transduction efficiencies as with targeting vectors. Also when the vectors were normalized to their p24 values, targeting vectors showed partial protection against α-MV antibodies in human plasma. Furthermore, the monoclonal neutralizing antibody K71 with a putative epitope close to the receptor binding sites of H, did not neutralize the targeting vectors, but did neutralize MV-LV. The observed escape from neutralization may be due to the point mutations in the H ectodomain that might have destroyed antibody binding sites. Furthermore, scFv mediated cell entry via the target receptor may proceed in presence of α-MV antibodies interfering with entry via the natural MV receptors. These results are promising for in vivo applications of targeting vectors in humans.Sabrina Kneissl; Tobias Abel; Anke Rasbach; Julia Brynza; Jürgen Schneider-Schaulies; Christian J. Buchholzarticlehttp://publikationen.ub.uni-frankfurt.de/frontdoor/index/index/docId/26759Thu, 11 Oct 2012 13:26:28 +0200